Genetically encoded infrared fluorescent protease reporters
AssigneeThe Regents of the University of California
InventorsXiaokun Shu, Tsz-Leung To
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Abstract
The invention described herein features infrared fluorescent protease reporters (iProteases) and methods of use thereof. The iProteases can be used in in vivo and in vitro assays to detect protease activity and disease states associated with protease activity. In a still further embodiment, the present invention provides a kit comprising any of the above described polynucleotides. In a further aspect, the present invention provides a method of in vivo optical imaging. In a still further embodiment, the in vivo imaging is performed in a living animal. In a further aspect, the present invention provides a method of detecting protease activity, the method comprising expressing a polypeptide according to any of those described above in a cell.
Claims19
A polypeptide comprising at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:24-46,
- wherein the polypeptide comprises an infrared fluorescent protein (IFP) PAS domain, an infrared fluorescent protein (IFP) GAF domain, a first split green fluorescent protein (GFP) domain, a second split green fluorescent protein (GFP) domain, and a protease cleavage site,
- wherein the first split GFP domain is attached to the IFP PAS domain, the second split GFP domain is attached to the IFP GAF domain and the IFP PAS and IFP GAF domains are joined by the protease cleavage site, and
- wherein infrared fluorescence is emitted upon cleavage of the protease cleavage site.
A method of detecting caspase mediated apoptosis, the method comprising expressing an infrared fluorescent protease reporter that has an amino acid sequence comprising at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs:40-42 in a cell, wherein the infrared fluorescent protease reporter fluoresces upon cleavage by caspase in response to caspase mediated apoptosis.
A method of detecting calpain activity, the method comprising expressing an infrared fluorescent protease reporter that has an amino acid sequence comprising at least 90% identity to amino acid sequence of SEQ ID NO:46 in a cell, wherein the infrared fluorescent protease reporter fluoresces upon cleavage by calpain.
Dependencies are inferred from the claim text.